The intestinal microflora is known to biotransform steroid hormones and bile acids into a variety of metabolites. Moreover, certain steroid metabolites generated by intestinal bacteria have been implicated as playing important roles in colon carcinogenesis, cholesterol homeostasis and gallstone formation in man. The overall objective of this proposed research is to delineate the role of intestinal bacterial steroid biotransformations on overall human steroid metabolism. The first goal will be to purify and characterize 21-dehydroxylase from the human intestinal anaerobe, Eubacterium lentum. The second goal will be to determine if human fecal suspensions and pure cultures of intestinal bacteria have C17-C20 lyase (desmolase) activity. The third goal will be to determine if intestinal bacteria can carry out 16 alpha-dehydroxylation of steroid hormones and efforts will be made to isolate bacteria responsible for this reaction. The fourth goal will be to purify and characterize the enzyme(s) involved in the 7 alpha- and beta-dehydroxylation of primary bile acids from Eubacterium sp. V.P.I. 12708. The final goal will be to isolate and biochemically characterize 7 alpha-dehydroxylase negative mutant strains of Eubacterium sp. V.P.I. 12708.